Assessing the risk factors associated with sarcopenia in patients with liver cirrhosis: a case–control study

Sarcopenia is a disease characterized by decreased muscle mass and strength, affecting 20–70% of patients with cirrhosis, and is associated with poor prognosis, complications, and high mortality. At present, the epidemiological investigation of sarcopenia in patients with liver cirrhosis is relatively limited, and because of the differences in population characteristics, regions, diagnostic criteria and diagnostic tools, the prevalence of sarcopenia in various studies varies greatly. The definition of sarcopenia in this study adopted the criteria of the Asian Working Group on Sarcopenia (AWGS 2019), including muscle mass and muscle strength / physical performance. A total of 271 patients with liver cirrhosis were included in this cross-sectional study to explore the influencing factors of sarcopenia in patients with liver cirrhosis. The prevalence of sarcopenia was 27.7%, 27.3% in male and 28.4% in female. The results of binary logistic regression analysis showed that age, physical activity, BMI, mid-upper arm muscle circumference, hepatic encephalopathy, nutritional status, alkaline phosphatase, albumin and total cholesterol were significantly correlated with the occurrence of sarcopenia in patients with liver cirrhosis. After adjusting for the potential influencing factors, it was found that the correlation between age and sarcopenia was weakened (OR = 0.870, 95% CI 0.338–2.239). The current findings show that sarcopenia is common in patients with cirrhosis and is independently associated with age, physical activity, BMI, nutritional status, and albumin, and serum alkaline phosphatase and total cholesterol are associated with the development of sarcopenia. Regular exercise may help maintain the grip strength of patients with cirrhosis and delay the deterioration of liver function.

The study was approved by the Medical Ethics Committee of the Affiliated Hospital of Zunyi Medical University (reference number: KLLY-2021-149).Written informed consent was obtained from all participants.All methods were carried out in accordance with relevant guidelines and regulations of the Affiliated Hospital of Zunyi Medical University Ethics Committee.

Variables and measurement
Outcome Sarcopenia was assessed as a dichotomous variable.According to the Asian Working Group for Sarcopenia 2019 (AWGS 2019), the diagnostic indicators of sarcopenia include three aspects: SMI, muscle strength, and physical mobility).The diagnosis of sarcopenia is made when the first criterion, plus the second and/or third criteria, are met 13 .

SMI
CT images were analyzed by two independent radiologists using the Slice-O-matic V5.0 software.Muscle tissues on CT images were based on Hounsfield unit (HU) thresholds, ranging from -29 HU to + 150 HU 14 .Skeletal muscle area (SMA) was measured by scanning the third lumbar spine vertebra cross-section, and the L3-SMI was calculated as the ratio of SMA to squared height (cm 2 /m 2 ).The cut-off point for low SMI was defined as SMI ≤ 52.4 cm 2 /m 2 for men and SMI ≤ 38.5 cm 2 /m 2 for women 15 .

Muscle strength
Hand grip strength (HGS) measurements were performed using an electronic hand grip strength device (EH 101, from Xiangshan, Guangdong, China) following the standardized grip strength measurement guidelines established by the American Manual Therapy Association 16 .Each participant was tested by maintaining a right angle (90°) to both hands, twice for both the right and left hands, using the average of the available maximum

Study size
The sample size was calculated using the formula: N = 4 ( U α S/δ) 2 , where U is the test level, U α represents the α corresponding U value, S represents the standard deviation, δ represents the tolerance error, and α=0.05.In this study, a two-sided test was performed with U = 1.96 and δ = [0.25S,0.5S] 5 .Considering the possible existence of invalid samples or data, the study increased the sample size by 20%.The calculated sample size range was approximately 75-296 cases, and 271 patients were finally included in this study.

Statistical analysis
Before any statistical comparison, the Kolmogorov-Smirnov test and homogeneity variance test were used to test the normality of standard normal distribution for all variables.In descriptive analysis, continuous variables were described with mean and standard deviation or median and interquartile range, whereas categorical variables were described using frequency and percentage.To indicate the association between independent risk factors and sarcopenia, the Student's t-test was used for continuous variables, and the χ 2 test was used for categorical variables.
The logistic regression model included the following covariates: age, physical activity, BMI, mid-upper arm muscle circumference, hepatic encephalopathy, nutritional status, alkaline phosphatase, albumin, and total cholesterol, and considered gender as a potential sarcopenia related factor.A three-step approach was employed to identify the risk factors associated with sarcopenia.First, risk factors for sarcopenia were evaluated and identified using simple regression analysis (i.e., Model 1).Second, adjusted binary regression was performed using a stepwise method.Three models were used to illustrate the association between these risk factors and sarcopenia: Model 1 represents the unadjusted model, Model 2 incorporates all significant risk factors identified using the stepwise method, and Model 3 represents the model adjusted to known confounding factors.

Institutional review board statement
This study protocol was approved by the Ethics Committee of the Affiliated Hospital of Zunyi Medical University on December 31, 2021, with the approval number: (KLLY-2021-149).

Informed consent statement
Informed consent was obtained from all subjects involved in the study.

Participants
A total of 482 hospitalized patients with LC initially met the inclusion criteria for the study and were selected for qualification screening.However, 201 of these patients met the exclusion criteria and were not eligible to participate.Of the eligible 281 cirrhotic patients, 10 of them were withdrawn due to physical discomfort during the screening process.Finally, 271 participants completed the study (Fig. 1).

Outcome data
Compared with participants without sarcopenia, subjects with combined sarcopenia had higher hospital costs, longer hospitalization periods (p < 0.001), and were more likely to develop spontaneous bacterial peritonitis, hepatic encephalopathy, ascites, and electrolyte disturbances (p < 0.05).Furthermore, the severity of liver function was strongly associated with the development of sarcopenia (p = 0.001) (Table 3).
The median hand grip strength was 22.8 kg for participants with sarcopenia and 28.9 kg for those without sarcopenia.Participants with sarcopenia had lower L3-SMI, L3-SMA, gait speed, total cholesterol, albumin, weight, BMI, and mid-arm muscle circumference than those without comorbidities (p < 0.001).Additionally, participants with sarcopenia had higher total bilirubin than those without sarcopenia (p < 0.001) (Tables 4&5).
Participants with sarcopenia had lower hemoglobin, triglycerides, high-density lipoprotein, and TSF than those www.nature.com/scientificreports/without comorbid sarcopenia (p < 0.05).Furthermore, participants with sarcopenia had higher gamma-glutamyl transferase and alkaline phosphatase than those without sarcopenia (p < 0.05).The incidence of sarcopenia in cirrhotic patients with high malnutrition risk was higher than that in patients with low-medium malnutrition (p < 0.05) (Table 5).

Outcome data
The binary ordered regression with sarcopenia as the dependent variable revealed several significant associations.In the adjusted model for the associated risk factors for sarcopenia (Model 2), the prevalence of sarcopenia increased with decreasing physical activity frequency (OR 2.498; 95% CI 1.063-5.874).High malnutrition risk was also linked to a higher prevalence of sarcopenia (OR 2.579; 95% CI

Discussion
In this study, hepatitis B infection was identified as the leading cause of cirrhosis, while alcoholic hepatitis was a common disease abroad.The prevalence of sarcopenia was higher in patients with cirrhosis (27.68%).In Model 1, the risk of sarcopenia was positively associated with age, physical activity < 3 times/week, high risk of malnutrition, hepatic encephalopathy, and high ALP levels.Conversely, it was negatively associated with high BMI, MAMC, albumin, and total cholesterol.After adjusting for gender in Model 3, the association between age and the risk of sarcopenia was attenuated.In addition, patients with advanced liver disease had a higher likelihood of developing sarcopenia.The prevalence of sarcopenia in patients with cirrhosis remains a subject of debate.A study that included 201 subjects with cirrhosis reported a prevalence of sarcopenia 57.2%, with no significant difference between males and females 19 .Kim et al. found that the complication rate of sarcopenia in patients with LC was 30%-70%, with a higher prevalence among males 20 .In the present study, there was no statistical difference in the prevalence of  www.nature.com/scientificreports/sarcopenia between males and females.This disparity in findings may be attributed to the differences in racial characteristics, body size, and disease etiology between individuals from Asian and Western countries.Notably, the prevalence of sarcopenia has been reported as 10% for Child-Pugh A, 34% for Child-Pugh B, and 54% for Child-Pugh C 21 , similar to the results of the present study, suggesting that muscle mass loss in cirrhotic patients is more pronounced as the liver reserve deteriorates.The present study made extensive efforts to account for potential confounding variables and identified aging as a significant risk factor for developing sarcopenia.However, this association was attenuated after adjusting for gender.This attenuation may be due to the small sample size of patients in the study.Research has shown that there can be a substantial loss of skeletal muscle mass, estimated at 20-30%, from ages 20 to 80 22 .Muscle strength begins to decline around the age of 30 and declines rapidly around the age of 50 years 23 .These age-related changes are characterized by motor unit remodeling, increased muscle fiber denervation, reduced protein synthesis, and a decrease in the number of muscle satellite cells required for skeletal muscle growth and repair 6 , ultimately leading to a decrease in muscle fiber cross-sectional area.Therefore, both the AWGS 2019 and the European Working Group on Sarcopenia in Older People 2 (EWGSOP 2) recommend early screening and intervention for sarcopenia in older adults to alleviate the burden of this condition in a high-risk population 1,13 .
Cirrhosis is usually associated with protein-energy malnutrition and low physical activity, leading to sarcopenia, with the prevalence of protein malnutrition ranging from 20 to 30% in patients with chronic liver disease (CLD) to over 60% in patients with cirrhosis 24 .Sarcopenia, characterized by decreased muscle/fat mass, increased pro-inflammatory cytokines, and anorexia, can potentially be mitigated with nutritional supplements, potentially slowing its progression to sarcopenia 25 .In the present study, a high risk of malnutrition (OR 2.579; 95% CI, 1.127-5.898)was associated with an increased risk of sarcopenia in cirrhotic patients with physical activity < 3 times/week (OR 2.498; 95% CI, 1.063-5.874).High ALP levels were also associated with the development of sarcopenia in cirrhotic patients.Lee et al. 26 explored the relationship between serum ALP levels and low muscle mass index in 15,579 adults in Korea, and the findings suggested that serum ALP may serve as a marker of inflammation and a predictor of sarcopenia.Further research is needed to confirm whether ALP can predict sarcopenia development in cirrhosis patients.Serum albumin levels have been recognized as an important factor Table 5.Comparison of laboratory and anthropometric indices between the sarcopenic group and the nonsarcopenic group..Montano et al. 28 also confirmed that decreased albumin levels were independently associated with a higher risk of sarcopenia and a poor prognosis.In the present study, albumin levels were significantly associated with the risk of sarcopenia in patients with cirrhosis.Regression analysis showed that normal total cholesterol levels were associated with the development of sarcopenia in cirrhotic patients.Previous studies have reported that obesity due to abnormal total cholesterol levels is a significant risk factor for the development of sarcopenia in cirrhotic patients 29 .BMI is an indicator to assess the nutritional status of the body.A low BMI typically indicates low body fat.Several studies have confirmed that L3-SMI is positively correlated with BMI 30,31 .In our study, the median BMI among participants with sarcopenia was 22.8 kg/m 2 , consistent with the findings of Landi 32 , who found that older people with a BMI greater than 21 kg/m 2 had 0.76 times the risk of sarcopenia compared to those with a BMI less than 21 kg/m 2 .MAMC is strongly associated with lean muscle mass and body fat, has good intraand inter-observer reproducibility, and has been shown to predict mortality 33,34 .While muscle mass is crucial, muscle strength is a more significant factor in determining physical function 35 .HGS has proven to be a better predictor of poor clinical outcomes than CT measurements of muscle mass and the Model for End-Stage Liver Disease.Even small improvements in HGS, as little as 1 kg, suggest a significant reduction in mortality 36,37 .Early identification of sarcopenia through routine imaging, convenient assessment tools, and clinical observation is an integral part of the care of patients with cirrhosis.Considering that screening tests need to be of high sensitivity, we recommend the combination of MAMC and grip strength as an initial screening test for sarcopenia rather than SARC-F in patients with CLD.SARC-F has demonstrated low sensitivity (15%-45%) in patients with CLD 38 , limiting its clinical application.Early detection of sarcopenia facilitates timely referral for confirmation of the diagnosis and implementation of primary prevention measures in high-risk populations.
This study confirms that hepatic encephalopathy is an independent risk factor for the development of sarcopenia.Hyperammonemia has been shown to increase muscle growth inhibitor expression through toll-like receptor-independent activation of nuclear factor kappa β in animal models 39 , increasing the risk of developing sarcopenia in patients with cirrhosis.Furthermore, Nardelli et al. 40 reported an association between the development of sarcopenia in patients with cirrhosis and the development of hepatic encephalopathy.Several studies have reported that branched-chain amino acids (BCAA) are beneficial in the treatment of hepatic encephalopathy 41 .For cirrhotic patients with protein intolerance or inability to meet protein targets, guidelines recommend a BCAA intake of 0.25 g•kg −1 d −142 .However, there is a paucity of research on the potential of BCAA in ameliorating sarcopenia.
The current study has several limitations.First, this is a case-control study conducted in a local hospital in China, which could potentially limit the generalizability of the findings.Second, there was no healthy control group to serve as a reference for comparing the prevalence of sarcopenia; therefore, only indirect comparisons could be made.Third, the sample size was relatively small, and there may be inherent bias among cases.In future

Conclusions
In conclusion, this study found a higher prevalence of sarcopenia in patients with cirrhosis.Advanced age, low physical activity, low BMI, decreased MAMC, hepatic encephalopathy, and nutritional status were associated with an increased risk of sarcopenia, whereas liver function-related indices such as ALP, albumin, and total cholesterol levels were associated with an increased risk of sarcopenia.These findings suggest that promoting physical activity in cirrhotic patients, with a focus on reducing body fat and enhancing cognitive function, plays a crucial role in preventing sarcopenia.

Figure 1 .
Figure 1.Flow chart showing study participant selection.

Table 1 .
Diagnostic criteria of sarcopenia.All statistical analyses were performed using the Statistical Package for Social Sciences (SPSS) 18.0, and all statistical tests were two-sided, with differences considered statistically significant at p < 0.05.

Table 2 .
Comparison of socio-demographic data between the sarcopenic group and the non-sarcopenic group.URR urban-rural resident.

Table 3 .
Comparison of disease-related information between the sarcopenic group and the non-sarcopenic group.IQR, interquartile range, HE hepatic encephalopathy, HRS hepatorenal syndrome, SBP spontaneous bacterial peritonitis.

Table 4 .
Comparison of muscle-related indicators between the sarcopenic group and the non-sarcopenic group.L3-SMI L3 skeletal muscle mass index, L3-SMA L3 skeletal muscle area, HGS hand grip strength.

Table 6 .
Logistic regression analysis of factors influencing the development of sarcopenia in patients with liver cirrhosis.Model 1 included exercise, BMI, MAMC, hepatic encephalopathy, nutritional status, ALP, albumin, and total cholesterol.Model 3 adjusted for gender based on Model 2. OR odds ratio, CI confidence interval, ref Reference group of the categorical variable.www.nature.com/scientificreports/research, we plan to address these limitations by including a larger cohort of cirrhotic patients and conducting follow-up assessments to enhance the study's reliability and generalizability.